The secretomotor action of choline on salivary secretion has been studied in the isolated perfused mandibular gland of the rat. Choline made up in substituted Ringer solutions (Na concentrations of 40, 70, or 100 mM) was an effective secretomotor agonist in the concentration range of 1-100 mM and evoked secretory responses comparable with those of acetylcholine (0.05-1.0 microM) administered at similar Na concentrations. Continuous infusion of choline, in contrast to acetylcholine, did not lead to a fall off in the secretory response (desensitization or tachyphylaxis) until the choline concentration used exceeded approximately 80 mM. The secretomotor action of choline (46 mM) was blocked completely and reversibly by atropine (5 microM) but not by propranolol (2.5 microM) and phentolamine (2.5 microM) administered together. When acetylcholine (1.0 or 0.05 microM) was infused on a background of choline stimulation, a small additional secretory response was elicited whenever the choline stimulus was submaximal, and, regardless of the choline concentration employed, addition of acetylcholine always induced tachyphylaxis. The results suggest that choline and acetylcholine act on the same population of muscarinic receptors, but it is unclear why choline does not evoke tachyphylaxis. The response to choline allows us to exclude a number of the possible causes of tachyphylaxis that have previously been considered, so that an excessive buildup of cytosolic free Ca now remains as the most likely cause of the phenomenon.